Adult normal mice are resistant to Dengue virus in general. The old method for isolation and in vivo titration involved intracranial inoculation in newborn mice that induce rapid death (neurovirulence).  see the old paper from Schlesinger and Frankel Am J Trop Med Hyg 1952, 1 p66-77.

I'm not sure that usage of immundeficient mice like AG129 might help you because of the cost..

From mosquito crushed in pbs buffer, filtered 0.22µM you may inoculated 10-40 µl in each new-born (or suckling mice from swiss strains for example)..  . a recent paper about StLouis encephalitis virus use this inoculation system (Rosa et al.  PLoS Negl Trop Dis. 2013 Nov 21;7(11):e2537. see the link below).

Usage of cell culture to amplify and isolate the virus should be considered ie C6/36 cells are highly sensitive even if the replication titer is not always very high and subsequent PCR may help to assess you system.

http://www.plosntds.org/article/info%3Adoi%2F10.1371%2Fjournal.pntd.0002537

Thank you so much. Please can you send the papers to [email protected] We have a problem of power in West Africa. I am considering using baby mice, precisely day old to amplify as we do not have C6/36 which is ideal.

There really aren't any good mouse models for Dengue virus. As Pierre suggested, the most common method is to use C6/36 cells to amplify the virus present in the mosquitoes. Essentially, homogenize the mosquito in L-15 media + 3% FBS (We use this percentage because any more causes the media to foam in our homogenizer). Once you have crude mosquito lysate, filter it through a 0.22um filter and inoculate a F-25 flask with a confluent monolayer of C6/36 cells. For our system we'll then incubate the flasks for 6 days, collect the media, replace it with fresh, and then collect is again at 8 days post-infection. This is usually enough to give us titers in the 10^7-10^8 ffu/ml range. From there you can do an RNA extraction from the supernatant and do RT-PCR to make cDNA. In my experience using this method, if I isolate RNA from 2ml of infected supernatant, I will recover RNA, but not enough to quantify. I just set up my RT-PCR with as much eluted RNA as I can, and the cDNA usually works just fine for PCR after that.

Good luck!

I'd like to concur with Pierre Andre Roques: intracranial inoculation of suckling mice. Our lab uses this method to screen for unknown arboviruses from mosquito pools and human specimens, as well as growing viruses for antigen preparations. We use 2-day old outbred NIH and inbred BALB/C strain mice but any lab strains will work; we have standardized on 2 day old mice because the heads of 1 day old mice are small and the cranial blood vessels are easily damaged while the crania of 3 day old mice (when the ears detach from the scalp) are beginning to ossify. For a dengue virus, allow 8-10 days or more for inoculated mice to start showing symptoms.

We also use C6/36 cells as well as the more susceptible AP61 line. Cell culture is most certainly more convenient and less controversial than infecting lab mice.

An alternative, much more sensitive method than mice or cells is mosquito inoculation. Aedes aegypti is ideal, being readily available in Africa, easily colonized and highly susceptible to the dengue viruses. The method for dengue viruses was pioneered by Rosen & Gubler in 1974 (file attached). I typically get titres in excess of 10 logs TCID50/ml with DEN-1 and DEN-2 viruses. However, one will need to ensure containment to avoid escape of infected mosquitoes and the technique does require a machined component and a way of drawing fine needles from glass capillaries.

Thanks Alan and Henri for your comments and because It seems that Allan like to have the "seminal" work.  here I include for Anyebe the 1952's paper and a more recent review from Indian collegues :-)

conventionally  mice was used for dengue virus isolation. In fact we tried in our laboratory. However, use of C6/36 is very convenient and easy to handle, since many arboviruses can be isolated in C6/36. It has been shown by many publications of Dr.A.B.Sudeep, Scientist from National Institute of Virology, Pune, India. 

Sorry everyone, have been having problem with my internet access. I really appreciate all the wonderful publications you have sent to me. They are great and very useful.

I agree with the protocol of Dr.Henri J Jupille.Similar way we are also follow for virus isolation.